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Involvement of eNAMPT/TLR4 signaling in murine radiation pneumonitis: protection by eNAMPT neutralization

      Therapeutic strategies to prevent or reduce the severity of radiation pneumonitis are a serious unmet need. We evaluated extracellular nicotinamide phosphoribosyltransferase (eNAMPT), a damage-associated molecular pattern protein (DAMP) and Toll-Like Receptor 4 (TLR4) ligand, as a therapeutic target in murine radiation pneumonitis. Radiation-induced murine and human NAMPT expression was assessed in vitro, in tissues (IHC, biochemistry, imaging), and in plasma. Wild type C57Bl6 mice (WT) and Nampt+/− heterozygous mice were exposed to 20Gy whole thoracic lung irradiation (WTLI) with or without weekly IP injection of IgG1 (control) or an eNAMPT-neutralizing polyclonal (pAb) or monoclonal antibody (mAb). BAL protein/cells and H&E staining were used to generate a WTLI severity score. Differentially-expressed genes (DEGs)/pathways were identified by RNA sequencing and bioinformatic analyses. Radiation exposure increases in vitro NAMPT expression in lung epithelium (NAMPT promoter activity) and NAMPT lung tissue expression in WTLI-exposed mice. Nampt+/− mice and eNAMPT pAb/mAb-treated mice exhibited significant histologic attenuation of WTLI-mediated lung injury with reduced levels of BAL protein and cells, and plasma levels of eNAMPT, IL-6,  and IL-1β. Genomic and biochemical studies from WTLI-exposed lung tissues highlighted dysregulation of NFkB/cytokine and MAP kinase signaling pathways which were rectified by eNAMPT mAb treatment. The eNAMPT/TLR4 pathway is essentially involved in radiation pathobiology with eNAMPT neutralization an effective therapeutic strategy to reduce the severity of radiation pneumonitis.

      Abbreviations:

      eNAMPT (extracellular nicotinamide phosphoribosyltransferase), DAMP (damage-associated molecular pattern protein), TLR4 (Toll-Like Receptor 4), WT (wild type), DEGs (differentially-regulated genes), WTLI (whole thoracic lung irradiation), BAL (bronchoalveolar lavage), mAb (monoclonal antibody), pAb (polyclonal antibody), EC (endothelial cells), HBE (human epithelial cells), MAPK (mitogen-activated protein kinase), ARDS (acute respiratory distress syndrome), PAH (pulmonary hypertension), IL-6 (Interleukin 6), IL-1 (Interleukin 1), RILI (radiation-induced lung injury), nmMLCK (non-muscle myosin light chain kinase), IR (ionizing radiation), 99mTc (Technetium), iQID (Quantum Imaging Detector), H&E (hematoxin and eosin), RILISS (Radiation induced lung injury severity score), ELISA (enzyme linked immunosorbent assay), FDR (False Discovery Rate), KEGG (Kyoto Encyclopedia of Genes and Genomes)
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